The long term objective of this proposal is to determine the molecular mechanisms that regulate cell division in E. coli and to apply this information to the study of other organisms. Recent work has focused on a cluster of cell division genes, ftsQ, ftsA and ftsZ, and has resulted in the determination of their nucleotide sequence. Utilizing this sequence information and a newly constructed, single copy, promoter detection vehicle the regulation of these genes will be further investigated. Of major interest is the regulation of these genes during the cell cycle and determination if these genes are autoregulated. Genetic evidence indicates that the FtsZ protein is the target of the SOS-inhibitor SulA which suggests that the FtsZ protein is at a key step in cell division. Overproduction of FtsZ results in increased cell division which is seen as the minicell phenotype. In addition, overproduction of FtsZ suppresses the UV-sensitivity of Ion by titrating out SulA. Future experiments are designed to look for SulA-FtsZ interaction by affinity chromatography, protein crosslinking and coimmunoprecipitation. These same experiments could also reveal interaction between FtsZ and other cellular proteins. These experiments are possible since FtsZ has been purified and antisera against FtsZ has been obtained. In addition, the cellular localization of FtsZ will be determined by immunoelectronnmicroscopy. Although the evidence is very strong that FtsZ plays a key role in cell division its biochemical function remains unknown. Recent analysis of the amino acid sequence revealed a region with strong homology to the DNA binding domain of repressor molecules. DNA cellulose chromatography will be used to examine the possibility of FtsZ being a DNA-binding protein. Also, additional mutations will be isolated in the FtsZ gene both in vivo and in vitro to examine structure-function relationships. Recent analysis of the evolution of the FtsZ protein revealed that it is present in all bacterial species and that the E. coli ftsZ gene functions among other members of the Enterbacteriaceae. The FtsZ gene will be isolated from both gram negative and gram positive species to further examine the evolutionary conservation of this gene and to examine structure-function relationships of the FtsZ protein.